Evolution and consequences of multidrug resistant ribosome

多重耐药核糖体的进化和后果

• 批准号: 10264066
• 负责人: Mee-Ngan F Yap
• 金额: $ 45.33万
• 依托单位: NORTHWESTERN UNIVERSITY AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-16 至 2025-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10264066
• 关键词: Address; Adenine; Affect; Antibiotics; Antimicrobial Resistance; Bacterial Infections; Binding; Biochemical; Biochemical Genetics; Biochemistry; Biological Assay; Carotenoids; Catalysis; Cells; Cessation of life; Ciprofloxacin; Clinical; Communities; Complement; Complex; Coupled; Cryoelectron Microscopy; Crystallization; DNA Gyrase; Data; Disulfides; Enzymatic Biochemistry; Enzymes; Evolution; Exhibits; Family; Gene Expression; Generations; Genes; Genetic Polymorphism; Genome; Goals; Gram-Positive Bacteria; Growth; Health Care Costs; Impairment; In Vitro; Infection; Knowledge; Laboratories; Link; Macrolides; Maps; Mass Spectrum Analysis; Measures; Mediating; Medical; Messenger RNA; Methyltransferase; Methyltransferase Gene; Mining; Modeling; Modification; Molecular; Multi-Drug Resistance; Mutation; Nucleotides; Operon; Pathway interactions; Peptides; Population; Protein Biosynthesis; Proteome; Proteomics; RNA; RNA Binding; Regulatory Pathway; Reporter; Resistance; Ribosomal RNA; Ribosome Inactivation; Ribosomes; Role; Site; Site-Directed Mutagenesis; Staphylococcus aureus; Streptogramins; Structure; System; Testing; Therapeutic; Toll-like receptors; Translations; Treatment Failure; Up-Regulation; Virulence; antimicrobial; bacterial genetics; bacterial resistance; base; comparative genomics; cost; crosslink; cytotoxic; design; experimental study; fitness; gel mobility shift assay; genetic approach; genome sequencing; global health; human disease; human pathogen; improved; in vivo; inhibitor/antagonist; innovation; insight; lincosamide; next generation sequencing; novel strategies; preference; public health relevance; resistance mechanism; ribosome profiling; structural biology; trait; translatome

ABSTRACT Posttranscriptional modifications of bacterial and eukaryotic ribosomes are linked to many human diseases, but the precise role of most modifications remains undefined. Dimethylation of a universally conserved adenine, A2058, in bacterial rRNA causes cross-resistance against all three critically important families of antibiotics (macrolides, lincosamides, and streptogramins (MLS)). A2058 dimethylation occludes MLS from the ribosome, thereby allowing normal protein biosynthesis and bacterial growth. The thirty-five classes of Erm methyltransferases responsible for A2058 dimethylation are invariantly encoded by a two-gene operon preceded by a short ribosome stalling leader sequence. These short stalling peptides considerably vary in size and sequence composition. The functional and evolutionary connections between the stalling sequence and its cognate erm gene are poorly understood. A previous `ribosome stalling' model suggests that macrolide-mediated translational stalling of the leader sequence is required for the upregulation of downstream co-transcribed erm, but clinical surveillance and our data indicate the existence of an alternative pathway. Our unpublished data further show that collateral sensitivity to unrelated antibiotics, reduction in virulence gene expression, accumulation of inactive ribosomes, and loss of in vivo fitness are all part of the trade-offs associated with the A2058 dimethylated ribosome. The exact mechanistic links between these traits are unknown. There is also an unmet need to understand the mechanism by which Erm recognizes and acts on 23S rRNA. This proposal will use a multi-pronged approach consisting of high-precision next-generation sequencing, bacterial genetics, proteomics, comparative genomics, biochemistry and structural biology to address three central questions: What are the underlying mechanisms of the trade-offs conferred by the A2058 dimethylated ribosome? How does the erm operon evolve, and how is the expression of erm regulated? How does Erm find its target substrate RNA? The erm operons are widespread among nosocomial Gram-negative and Gram-positive bacteria, addressing these questions will offer significant mechanistic insight into new antimicrobial strategies tailored to disrupt these biochemical interactions and regulatory pathways.

摘要 细菌和真核细胞核糖体的转录后修饰与许多人类 疾病，但大多数修饰的确切作用仍未确定。一种普遍存在的二甲基 细菌rRNA中保守的腺嘌呤A2058可引起对这三种病毒的交叉耐药 重要的抗生素家族(大环内酯类、林可酰胺类和链霉素类(MLS))。A2058 双甲基化封闭了核糖体上的MLs，从而允许正常的蛋白质生物合成和 细菌生长。与A2058二甲基化有关的35种ERM甲基转移酶 总是由一个双基因操纵子编码，前面有一个短的核糖体停滞前导序列。 这些短失速多肽在大小和序列组成上有很大的差异。功能和 人们对失速序列与其同源erm基因之间的进化联系知之甚少。 先前的核糖体停滞模型表明，大环内酯介导的领导者的翻译停滞 下游共转录的erm的上调需要序列，但临床监测和 我们的数据表明存在另一种途径。我们未公布的数据进一步表明， 对无关抗生素的间接敏感性，毒力基因表达的减少， 不活跃的核糖体和体内适应性的丧失都是与A2058相关的权衡的一部分 二甲基化核糖体。这些特征之间的确切机制联系尚不清楚。还有一个 未满足的需要了解ERm识别23S rRNA并作用于23S rRNA的机制。这 提案将使用由高精度下一代测序组成的多管齐下的方法， 细菌遗传学、蛋白质组学、比较基因组学、生物化学和结构生物学 三个核心问题：A2058实现权衡的潜在机制是什么 二甲基化核糖体？Erm操纵子是如何进化的，erm的表达又是如何调控的？ ERM是如何找到其目标底物RNA的？ERM操纵子在医院中广泛存在 革兰氏阴性和革兰氏阳性细菌，解决这些问题将提供重要的机制 洞察新的抗菌策略，以破坏这些生化相互作用和调控 小路。